Diabetes 45:1037-1043, 1996
© 1996 by the American Diabetes Association, Inc.
Advanced glycation end products of the Maillard reaction in aortic pepsin-insoluble and pepsin-soluble collagen from diabetic rats
ABSTRACT
Recent immunohistological studies using antibodies against advanced
glycation end products (AGEs) have demonstrated the presence of AGEs in
several tissues. By an enzyme-linked immunosorbent assay using the
monoclonal anti-AGE antibody, the present study aimed to determine AGEs in
pepsin-insoluble collagen (PIC) as well as in pepsin-soluble collagen (PSC)
from the aortas of streptozotocin (STZ)-induced diabetic rats (at 4, 16,
and 28 weeks after STZ injection) and those of age-matched control rats.
Addition of EDTA to the immunoassay buffer has led us to successful
determination of AGEs in the aortic PIC samples with following results: 1)
in diabetic rats, there was a time-related increase in the AGE contents at
28 weeks (n = 9, 226.4 +/- 13.5 ng/mg collagen [mean +/- SE]), compared
with that at 4 and 16 weeks (n = 6, 79.6 +/- 9.5 ng/mg collagen, and n = 8,
149.4 +/- 30.9 ng/mg collagen at 4 and 16 weeks, respectively; both P <
0.05, between 4 and 16 weeks and 28 weeks); 2) after 28 weeks of diabetes,
the AGE contents in PIC of aortas were significantly higher in diabetic
rats than in controls (n = 9, 226.4 +/- 13.5 ng/mg collagen vs. n = 8,
129.6 +/- 14.9 ng/mg collagen, P < 0.01, diabetic vs. control); and 3)
the level of the AGE content was strongly correlated with the PIC/total
collagen (TC) ratio (n = 45, r = 0.698, P = 0.0001). By treating the
samples of PSC with alkaline solution, the AGE content of PSC was also
determined. In the PSC fraction, the AGE levels in the diabetic rats tended
to increase with time and to be higher than those of control rats at 28
weeks although these changes were not statistically significant (diabetic:
n = 4, 19.4 +/- 9.7; n = 6, 22.3 +/- 6.2; n = 6, 39.6 +/- 10.8; control: n
= 4, 19.7 +/- 9.8; n = 6, 22.9 +/- 7.3; n = 7, 30.7 +/- 7.2; at 4, 16, and
28 weeks, respectively). Compared with the AGE levels of PSC, those of PIC
were about four to seven times and four to five times higher in diabetic
and control rats, respectively (PIC versus PSC in diabetic or control rats,
all P < 0.001, at 4, 16, and 28 weeks, respectively). These findings
provide the first immunochemical evidence that AGE adducts are present in
the materials extracted sequentially by pepsin and collagenase and that
these adducts in PIC accumulated as a function of the increase in the
aortic PIC/TC ratio.

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Copyright © 1996 by the American Diabetes Association.
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